The present invention is directed to the field of immunology. In particular, the present invention is directed to the use of genetically-modified professional antigen-presenting cells to combat infections or tumors.
Critical analysis of early events in the cellular immune response to tumor and viral antigen have identified dendritic cells as the major antigen-presenting cells (APCs) eliciting an effective T cell response (Steinman (1991), Annu. Rev. Immunol. 9:271-296; Macatonia et al. (1989), J. Exp. Med. 169:1255-1264). Dendritic cells (DCs), appropriately activated, take up soluble antigen and apoptotic bodies, migrate to the paracortical T cell-rich areas of lymph nodes, and initiate a series of interactions leading to the selection of antigen-specific T cells, and to the release of the DC cytokines, interferon-xcex1 (IFN-xcex1) and interleukin-12 (IL-12).
It has previously been demonstrated that administration of DCs pulsed with synthetic tumor-associated peptides serve as effective therapeutic antitumor vaccines, inducing an effective antitumor immune response in vitro and following adoptive transfer in mice (Mayordomo et al. (1995), Nature Med. 1:1297-1302; Zitvogel et al. (1996), J. Exp. Med. 183:87-97; Porgador and Gilboa (1995), J. Exp. Med. 182:255-260; Porgador et al. (1996), J. Immunol. 156:2918-2926). However, T cell-defined epitopes have been identified only for a limited number of human tumor types. Several approaches to overcome this problem including pulsing DCs with acid-eluted bulk tumor peptides Zitvogel et al. (1996), tumor extracts and RNA (Flamand et al. (1994), Eur. J. Immunol. 24:605-610; Ashley et al. (1997), J. Exp. Med. 186:1177-1182; Boczkowski et al. (1996), J. Exp. Med. 184:465-472) or fusion of tumor with DC (Gong et al. (1997) Nature Med. 3:558-561) have been employed for DC-based vaccination strategies against tumors. Even though these approaches will allow treatment of tumors for which tumor associated antigen is not well characterized, there are still significant problems, particularly in the preparation of clinical samples from human solid cancers.
IL-12 is a heterodimeric cytokine produced by DCs, macrophages, polymorphonuclear leukocytes and keratinocytes (Lamont and Adorini (1996), Immnunol. Today 17:214-217). IL-12 enhances natural killer (NK) cell and cytotoxic T lymphocyte (CTL) activities, plays a key role in the induction of Th1 immune responses including IFN-xcex3 production, and has IFN-xcex3/interferon-inducible protein 10 (IP-10)-dependent antiangiogenic effects (Lamont and Adorini (1996), supra; Voest et al. (1995), J. Natl. Cancer Inst. 87:581-586; Sgadari et al. (1996), Blood 87:3877-3882). DCs are capable of producing IL-12 after ligation of CD40 and class II molecules, presumably only following interaction with T cells, and IL-12 delivery in conjunction with DCs enhances CTL response in vitro (Heufler et al. (1996), Eur. J. Immunol. 26:659-668; Koch et al. (1996), J. Exp. Med. 184:741-746; Bhardwaj et al. (1996), J. Clin. Invest. 98:715-722).
There have been reports of potent antitumor effects of IL-12 in a vaccination model with IL-12 gene-modified tumor cells, as well as with systemic administration of IL-12 protein (Brunda et al. (1993), J. Exp. Med. 178:1223-1230; Nastala et al. (1994), J. Immunol. 153:1697-1706; Tahara et al. (1995), J. Immunol. 154:6466-6474; Martinotti et al. (1995), Eur. J. Immunol. 25:137-146). Direct injection of IL-12-transduced fibroblasts also effectively eliminated established tumors with concomitant induction of effective systemic immunity (Zitvogel et al. (1995), J. Immunol. 155:1393-1403). Based on these results, an initial clinical trial of IL-12 gene therapy has been completed using autologous fibroblasts in the context of a phase I study (Tahara et al. (1997), Proc. Am. Soc. Clin. Oncol. 16:439a). Partial responses were observed in patients with melanoma, breast cancer, and head and neck tumors persisting for up to two years.
The present invention depends, in part, upon the discovery that professional antigen-presenting cells (APCs) which have been genetically modified to enhance expression of an immunostimulatory cytokine, may be directly injected into or near the site of an infection or tumor to induce a specific immunological response against antigens associated with the infection or tumor without pre-loading or pulsing the APCs with the antigens. In particular, it has been found that dendritic cells (DCs), and preferably bone marrow-derived dendritic cells (BM-DCs) or CD34+-derived dendritic cells (CD34+-DCs), which have been genetically modified to enhance expression an immunostimulatory cytokine, preferably interleukin-12 (IL-12), may be injected into or near the site of an infection or tumor to induce a specific immune response against antigens associated with the site of injection.
Therefore, in one aspect, the present invention provides methods for treating an individual having an infection or tumor comprising injecting the individual near or at the site of the infection or tumor with an effective amount of professional antigen presenting cells (APCs) which have been genetically modified to enhance the expression of an immunostimulatory cytokine.
In preferred embodiments, the genetically modified APCs are professional antigen presenting cells, and most preferably the PAPCs are dendritic cells selected from the group consisting of CD34+-derived dendritic cells, bone marrow-derived dendritic cells, monocyte-derived dendritic cells, splenocyte derived dendritic cells, skin-derived dendritic cells, follicular dendritic cells, and germinal center dendritic cells. In particularly preferred embodiments, the dendritic cells are CD34+-derived dendritic cells cultured in the presence of at least one factor selected from the group consisting of G-CSF, GM-CSF, TNF-xcex1, IL-4, the Flt-3 ligand and the kit ligand.
In addition, in preferred embodiments, the immunostimulatory cytokine is selected from the group consisting of the interleukins (e.g., IL-1xcex1, IL-1xcex2, IL-2, IL-3, IL-4, IL-6, IL-8, IL-9, IL-10, IL-12, IL-18, IL-19, IL-20), the interferons (e.g., IFN-xcex1, IFN-xcex2, IFN-xcex3), tumor necrosis factor (TNF), transforming growth factor-xcex2 (TGF-xcex2), granulocyte colony stimulating factor (G-CSF), macrophage colony stimulating factor (M-CSF), granulocyte-macrophage colony stimulating factor (GM-CSF), the Flt-3 ligand and the kit ligand.
Further, in preferred embodiments, the APCs have been genetically modified by transduction with a viral vector encoding the immunostimulatory said cytokine and, most preferably, with a retroviral vector. In other embodiments, however, the APCs may be genetically modified with adenoviral vectors or adeno-associated viral vectors, or by lipofection, ballistic injection, or other means of genetic modification known in the art.
Further, in any of the foregoing embodiments, the individual which is treated may suffer from a cancer selected from the group consisting of melanomas, hepatomas, adenocarcinomas, basal cell cancers, oral cancers, nasopharyngeal cancers, laryngeal cancers, bladder cancers, head and neck cancers, renal cell cancers, pancreatic cancers, pulmonary cancers, cervical cancers, ovarian cancers, esophageal cancers, gastric cancers, prostate cancers, testicular cancers, breast cancers, or other solid tumors. Alternatively, the individual may suffer from a refractory infection.